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EMEA Workshop on Biosimilar Monoclonal Antibodies, July 2, 2009
Session 1 CMC
Innovator Industry Presentation
Prof. Georg-B. Kresse
London July 2, 2009
G.-B. Kresse / CMC
1
The Innovators‘ View – CMC Summary
• Available guidance for quality characterization is applicable for
biosimilar mAbs
• The mode of action of mAbs is complex and may involve contributions
from multiple mechanisms
• Differences in antibody variants are only acceptable if justified by clinical
data
• Glycosylation can be critical for the biological function of mAbs
• Quality data cannot substitute for gaps in knowledge in functional
assays
• Role of ICH Q8 and Q9: a „design space“ concept is not transferable
from the reference product to a biosimilar to ensure similarity
London July 2, 2009
G.-B. Kresse / CMC
2
The EU Biosimilars Existing Regulatory Framework
• It is impossible to characterize the quality attributes of proteins completely by
physicochemical analysis, and to fully predict the impact of structural
differences (induced by the producing cell itself as well as by the manufacturing
process) on clinical efficacy and safety.
• For independently manufactured protein products, „identical copies“ are
impossible but at best „similar“ products. Biosimilars are manufactured and
controlled according to their own development. Similarity has to be shown in
terms of quality, efficacy, and safety in head-to-head comparative studies.
• Analytical differences between
biosimilars and their reference
product are expected. The extent of
observed differences will be decisive
for definition of the non-clinical and
clinical program.
• The „biosimilarity“ scenario differs
from the „comparability after
manufacturing changes“ scenario
regulated by ICH Q5E.
Source: P. Richardson and P. Celis (2007), 1st Drug Evaluation Forum, Pharm. Soc. Japan
London July 2, 2009
G.-B. Kresse / CMC
3
Available Guidance for Quality Characterization
is applicable for biosimilar mAbs
• The same principles should apply for quality characterization of biosimilar
mAbs as for other biosimilars.
• Available guidelines on Quality Characterization of mAbs (CHMP/BWP/157653/2007)
and on Quality Issues of Biosimilar Products (CHMP/BWP/49348/2005) are applicable
for biosimilar mAbs. There is no need for additional quality guidance.
• State-of-the-art analytical methods will have to be used. However, with more
sensitive analytics, more differences will be detected. On the other hand, even
extended analytical characterization is limited – „you will only see what you are
looking for“.
• Both physicochemical and biological assays will be necessary in all cases.
• The quality studies have to take into account multifunctionality of mAb
molecules and should include in-vitro potency assays (if predictive assays are
available) as well as assays for Fab and Fc mediated functions as needed based
on an understanding of the mechanism of action of the particular mAb.
Assumptions based e.g. on the IgG sub-class are not sufficient.
London July 2, 2009
G.-B. Kresse / CMC
4
The Mode of Action of mAbs is complex
and may involve Contributions from multiple Mechanisms
Complement
C1q
Activation (CDC)
(Example: Rituximab)
3
CD
R2
HE
HE R2
FR
G
E
CD
20
Induction of Apoptosis
(Example: Rituximab)
G
VE
R
F-
T-Cell
CD20
Tumor
Cell
R2
NK-Cell
CD
20
HE
Activation of Effector
Mechanisms
Antibody-dependent cellular
Cytotoxicity (ADCC)
(Example: Rituximab)
Inhibition of Signal Transduction or Receptor Activation
• Inhibition of Ligand Binding (Example: Cetuximab)
• Induction of Receptor Internalization (Example: IGF-1R-Abs)
• Inhibition of Receptor Dimerization (Example: Pertuzumab)
• Inhibition of Receptor Shedding (Example: Trastuzumab)
Activation of T-Cells
(Example: Catumaxomab)
Targeting of Toxins
(Example: T-DM1)
Blocking Ligand Binding
(Example: Bevacizumab)
The in-vivo net contribution of different modes of action described for one mAb is
often incompletely understood and may also be different in different indications.
London July 2, 2009
G.-B. Kresse / CMC
Modified from: Hasmann, M. et al. (2009) ChiuZ, submitted
5
Differences in Antibody Variants are only acceptable
if justified by clinical Data
• Biosimilars must have the same amino
acid sequence as the reference product.
• The relevance of major variants (e.g.,
basic/acidic variants, presence or absence of
C-terminal Lys residues) on clinical efficacy
and/or safety has to be established and
reflected in the control strategy of the product.
• mAbs (reference and biosimilar) will always
be micro-heterogeous mixtures of a large
number of post-translationally modified
molecular species.
• The exact composition of this mixture cannot
be reproduced if a different manufacturing
process is used, therefore comparative nonclinical and clinical data will always be
necessary for biosimilar mAbs.
London July 2, 2009
G.-B. Kresse / CMC
Not all possible variants are described. For example,
there are fucosylation variants in glycosylation that
were not counted. If one assumes these variants are
independent and considers combinations, each halfantibody has 2x6x4x4x5x5x2=9600 possible states.
If one assumes both halves of the antibody are
independent, there are (9600)2 ≈ 108
possible
states.
Source: Kozlowski, S. & Swann, P. (2006) Adv. Drug
Delivery Revs 58, 707-722
6
Glycosylation can be critical for the biological Function of mAbs
• IgGs may contain up to ~500 different glycoforms
due to Fc glycosylation. Differences may influence
solubility, stability, clearance, immunogenicity, and
immune effector functions.
• Even small differences in glycosylation may have
significant effects (e.g., impact of absence of one
fucose residue on ADCC - however, not only
fucosylation is important).
Source: Jefferis, R. (2009) Nature Revs.
Drug Disc. 8, 226-234
• Up to 30% of human IgGs contain N-linked
oligosaccharides in the Fab region whose functional
significance is not fully evaluated (e.g., impact of Fab
galactosylation on hypersensitivity reaction).
• The pattern of glycosylation will vary between products because it depends on
the manufacturing process.
• In
line with the mAb guidance, the relevance and acceptability of glycosylation
differences should depend on criticality of this attribute, i.e. the proven or
disproven impact of glycosylation differences on clinical properties which may
be different for mAbs using different modes of action.
London July 2, 2009
G.-B. Kresse / CMC
7
Need for functional Assays Quality Data cannot substitute for Gaps in Knowledge
• The broad experience existing with
therapeutic mAbs shows that it may be
difficult to understand the critical
quality attributes, and to predict the
impact of differences to clinical
efficacy and safety (e.g., Raptiva
example).
Direct Effects mediated
by Fab Part
Target cell
• Only those differences known/proven
to have no impact on clinical efficacy
and/or safety should be acceptable
without additional justification.
FcR
CDC
Immune
Effector
Effector
Functions
cell
mediated
by Fc
Interactions
ADCC
• Gaps of functional knowledge will lead to the requirement for additional nonclinical and clinical data, based on knowledge of the mode of action of the
particular mAb under study.
• Because of the linkage of quality, non-clinical and clinical aspects, a „holistic
approach“ is needed for the evaluation of mAb-based drugs to connect
analytical data with clinical safety and efficacy.
London July 2, 2009
G.-B. Kresse / CMC
8
Role of ICH Q8 and Q9: A „Design Space“ Concept is not
applicable to ensure Similarity
• ICH Q8 Pharmaceutical Development and ICH Q9 Quality Risk Management
are applicable for biosimilars manufacturers for their own developments in the
same way as for the originators.
• A „design space“ depends on a particular manufacturing process connected to
clinical results and cannot be „borrowed“ to demonstrate similarity of a biosimilar
product to a reference product made by a different process.
Analyze product quality
attributes and batch-to-batch
variability
No access to proprietary data of innovator.
Design Space will be different for products
manufactured differently.
Understand relevance / impact
to clinical safety and efficacy
Batch-dependent clinical data of reference
product are not available for a second
manufacturer
Define design space
Define Control Strategy
London July 2, 2009
Design space of reference product cannot be
utilized by a second manufacturer
Second manufacturer needs to establish own
control strategy based on own data.
G.-B. Kresse / CMC
9
CMC Conclusion
• Available quality guidance is applicable for biosimilar mAbs.
• It is impossible to characterize the quality attributes of mAbs completely
by physico-chemical analysis alone and to fully predict the impact of
differences on clinical efficacy and safety. Analytical differences (including
glycosylation if appropriate) have to be justified by clinical data.
• Multifunctionality of mAb molecules means that characterization should
include both Fab and Fc mediated functions unless justified.
• The mode of Action of mAbs is often complex involving contributions from
multiple mechanisms (these may be different in different indications).
• mAbs will always be micro-heterogeneous mixtures of a large number of
molecular species. The relevance of major variants on clinical efficacy and/or
safety has to be established and reflected in the control strategy of the product.
• ICH Q8 and Q9 are applicable for biosimilars manufacturers for their own
developments as for originators. However, a „design space“ concept will not help
to establish similarity.
London July 2, 2009
G.-B. Kresse / CMC
10
Back-up Slides
London July 2, 2009
G.-B. Kresse / CMC
11
Manufacturing of recombinant Proteins is complex
DNA Vector
Cloning into
DNA Vector
Transfer into Host Cell
Expression
Cell Development
Downstreaming
Large-Scale Fermentation
e.g., bacterial or mammalian cell
London July 2, 2009
Formulation
G.-B. Kresse / CMC
12
Manufacturing of recombinant Proteins is complex
A second manufacturer uses...
DNA Vector
(Probably)
a different
DNA vector
The same amino
acid sequence
Cloning into
DNA Vector
Transfer into Host Cell
Expression
Cell Development
(maybe the same genetic
sequence)
A different
fermentation
process
A different
downstreaming
protocol
Downstreaming
Different
in-process
controls
A different
recombinant cell
expression
e.g., bacterial
or mammaliansystem
cell
Large-Scale Fermentation
Formulation
Different Process → different product
London July 2, 2009
G.-B. Kresse / CMC
Maybe a
different
formulation
13
Are mAbs „well-characterized Biologicals“ ?
• The term „well-characterized biological“ should not be used because is not
defined in any EMEA guideline.
• It was originally introduced by FDA in 1995 but soon abandoned:
In the Federal Register of December 8, 1995 (60 FR 63048), the agency first published an
interim definition of a well-characterized therapeutic recombinant DNA-derived and
monoclonal antibody biotechnology Product … After considering the public comments
received on the interim definition … and the many requests the agency has received for further
clarification of the term ‘‘well-characterized,’’ FDA has determined that it may not be possible
to achieve a sufficiently clear and specific understanding of this term to adequately apprise
potential applicants of the applicability of the new procedures. Accordingly, FDA is specifying,
in lieu of the term ‘‘well characterized biotechnology product,’’ the categories of products to
which this final rule will be applicable.” (Federal Register / Vol. 61, No. 94 / Tuesday, May 14,
1996)
London July 2, 2009
G.-B. Kresse / CMC
14
Source: P. Richardson and P. Celis (2007), 1st Drug Evaluation Forum, Pharm. Soc. Japan
London July 2, 2009
G.-B. Kresse / CMC
15
Standard for documenting Innovator Process Changes is
well established – Standard for Biosimilars should be no less
2nd generation innovator processes – and Biosimilars
Appropriate human study to demonstrate the equivalence of the
efficacy and safety (including immunogenicity) of the product
Biosimilars
Manufacturing Change
Change
filter
supplier
Move
equipment
different part
of facility
• Low risk
• Frequent
• Supported by:
Analytical and
Process Data
Move
Scale-up
New cell line
manufacturing Manufacturing
New Process
to new facility
New Formulation
Character of Change
• Highest risk
• Rare
• Extensive Data:
Analytical, Process
and Human data
New:
• DNA ?
• Cell line
• Process Technology ?
• Fermentation process
• Purification process
• Analytics
• Facilities
• Formulation ?
• And - no history
Source: modified from Ken Seamon, Cambridge University
London July 2, 2009
G.-B. Kresse / CMC
16
Second Manufacturers have no Access
to the Innovator‘s Database
Technical Development
Clinical Development
Drug Product
No access to
Second
Manufacturer historical database
Limited access to
historical database
Comparison at
the level of final
product only
(limited to drug
product level)
Innovator
No definition of
“design space” and
assessment of
similarity possible
Case-by-case: clinical
development can be
shortened
Trade secret of innovator
Accessible for independent second manufacturer
London July 2, 2009
G.-B. Kresse / CMC
17
Process Control
• Procedures
• Materials
• In-process testing
• Monitoring
• Validation
End-Product only
In-Process
Controls
(Process & Product)
Comparability
Extended
Characterization
Similarity
Release Tests
(Specifications)
Consistency
How much of the Iceberg is visible?
Unknown
Learned over time –
update control strategy
London July 2, 2009
Modified from: Koszlowski, S. & Swann, P. (2006) Adv. Drug Delivery Revs. 58, 707-722
G.-B. Kresse / CMC
18
Is physicochemical Characterization sensitive enough to detect
Differences between two more complex Molecules like mAbs ?
• Sensitivity is not the issue – the analytical toolbox becomes more and more
comprehensive, and state-of-the-art analytical methods have to be used by biosimilars
developers as well as by innovators.
• With more sensivity analytics, more differences will be detected - not „more identity“.
„The greater the resolution of the analytical methods applied to
a product, the more heterogeneity will be apparent. The critical
issue is which variants matter and at what levels they matter“
Kozlowski, S. & Swann, P. (2006) Adv. Drug Delivery Revs. 58, 707-722
• According to the biosimilars quality guideline, differences are expected anyway –
but the extent of observed differences decisive for definition of the non-clinical and
clinical program.
It is not expected that the quality attributes in the similar biological and reference medicinal
products will be identical. For example, minor structural differences in the active substance, such
as variability in post-translational modifications may be acceptable, however, must be justified. …
Therefore, differences … may have consequences with regard to the amount of non-clinical
and clinical data which may be required in order to make satisfactory justification of the safety
and efficacy of the similar biological medicinal product.
EMEA/CHMP/49348/05
London July 2, 2009
G.-B. Kresse / CMC
19
Even small Glycosylation Differences may have
significant Effects on Immune Effector Functions
• Both amino acid sequence and glycosylation
pattern of CH2 influence FcR binding and ADCC
activity.
• The presence or absence of one fucose residue
can affect the biological activity (killing of target cells
via ADCC).
• Even very small differences in fucosylation may
have significant effects on in vitro ADCC.
Immune effector function
cell lysis in %
120
glycoengineered antibody
100
(only 26% fucosylated)
80
wildtype antibody
60
(100% fucosylated)
40
glycoengineered negative
control antibody
20
Clark, M., http://wwwimmuno.path.cam.ac.uk/~mrc7/
0
10
20
30
40
50
antibody conc. [ng/ml]
Source: Roche
London July 2, 2009
Source: R. Garnick (2008)
Biogenerics 2008 Conference
Relative ADCC (in-vitro)
0
5
4
3
2
1
0
0
1
G.-B. Kresse / CMC
2
3
4
% Non-Fucosylated IgG
5
6
7
20
Not only Fucosylation is important
GlcNAc/
Mannose
Ligand for Mannose Binding Protein Æ
complement activation (Malhotra et al., Nat.
Med. 1995)
Sialic acid
Suppression of ADCC (anti-inflammatory
activity) (Kaneko et al., Science 2006)
Galactose
Placental transport
(Kibe et al., J. Clin. Biochem. Nutr. 1996)
bisecting
GlcNAc
Prevents core fucosylation Æ enhanced
ADCC (Umaña et al., Nat. Biotech. 1999)
absence of
core Fucose
Enhanced ADCC
(Okazaki et al., J. Mol. Biol. 2004)
α(1-3)-Gal
a1-3
Non-human/antigenic
(Cooper, Xenotransplantation 1998)
Source: H. Kettenberger, Roche
London July 2, 2009
G.-B. Kresse / CMC
21
Correlation of Bioactivity and Galactose Content in Rituximab
In Vitro CDC Bioactivity
120
110
100
90
80
β- galactosidase treated
70
60
50
0
0.1
0.2
0.3
0.4
0.5
0.6
Moles Galactose per Mole Heavy Chain
0.7
0.8
Source: Genentech, Inc.
London July 2, 2009
G.-B. Kresse / CMC
22
The Glycopattern depends on the Host Cell
and on the Fermentation Conditions
Level of a-fucose during 10-l-lab scale fermentation of a glycoengineered antibody
Clone 1, 1 X GF
Run 5 #165
% non-fuc
Run 2.1
Run 2.3
40%
40%
20%
20%
20%
0%
0%
0%
0%
9
7
17
15
13
11
9
7
5
15
14
13
12
11
10
9
8
11
Run 2.4
Run 2.2
7
17
40%
20%
6
60%
5
60%
15
60%
40%
14
60%
13
80%
12
80%
11
80%
10
80%
9
100%
8
100%
7
100%
17
5
15
15
14
13
12
11
9
10
8
7
6
5
100%
6
Run 6 #165
5
Run 2 #1
15
0%
15
0%
13
0%
13
0%
11
20%
17
20%
15
20%
13
20%
11
40%
9
40%
7
40%
5
40%
14
60%
13
60%
12
60%
11
60%
9
80%
10
80%
8
80%
7
100%
80%
6
100%
5
100%
% bisected
9
% complex
100%
7
% no n-fuc
% co mp lex
% b isected
5
Run 1 #1
Clone 165, 1 X GF Clone 165, 1 X GF Clone 165, 2 X GF
Source: Roche
London July 2, 2009
G.-B. Kresse / CMC
23
Up to 30% of native human IgGs contain Glycosylation
in variable Regions of LC and HC
• Mostly not germline encoded; created by somatic mutation. May also be created
in mAbs obtained by phage display technology.
• Occupation ranges from (almost) complete to low percent, depending on
glycosylation sequon, accessibility, and process conditions (expression system,
cell line, fermentation process).
• Different glycan types may be present → microheterogeneity
• Potentially has significant influence on functional and pharmacological
characteristics (e.g., ligand binding, stickyness/unspecific binding, clearance
rate, and potency) and technical development (consistency of isoform mixture,
pK value, stability, comparability assessment).
• If Fab glycosylation sites are present
in the reference mAb, additional effort
required for analytical characterization,
documentation and reporting.
London July 2, 2009
G.-B. Kresse / CMC
24
The Relevance of Fab Glycosylation:
The Cetuximab Example
C.H. Chung et al. (2008) NEJM 358, 1109-1117
London July 2, 2009
G.-B. Kresse / CMC
25
The Efalizumab Experience
• Raptiva (efalizumab) was originally manufactured by XOMA and used in the Phase I/II trials
and up to Phase III. Manufacturing was transferred to Genentech with full information.
Analytical and formulation differences expected to be inconsequential were observed.
• In a bioequivalence study, differences
in PK parameters were observed:
AUCinf
27.9
GNE
36.9
Ratio
(G:X)
Percentage of Subjects
Xoma
• A trend for lower PASI* Response to
Efalizumab was found despite higher peripheral
drug concentrations
1.324
AUCt
26.9
35.6
1.324
Cmax
3.59
4.22
1.175
• Based on the PK data, one could imagine
that administering ~70% of the dose of the
Genentech antibody would have similar
effects to the XOMA antibody. However,
because of the unpredictable nature of
these PK changes an additional Phase III
study was performed to determine the safety
and efficacy of the Genentech material.
London July 2, 2009
45%
40%
35%
30%
25%
20%
15%
10%
5%
0%
38,9%
improved 75%
26,6%
*PASI = Psoriasis
Area and Severity
Index
2,4%
Placebo
(n=170)
1.0 mg/kg/wk 1.0 mg/kg/wk
GNE
XOMA
material
material
(n=369)
(n=162)
Source: Genentech
Changes that were believed not to affect the
properties of the protein resulted in clear
differences in pharmacokinetics.
Given the complexity of therapeutic proteins, the
impact of changes in pharmacokinetics (and
probably pharmacodynamics) on safety and
efficacy can often not be predicted reliably.
G.-B. Kresse / CMC
26
Compendial Methods may fail to reveal Variants
with changed Biopotency and Receptor Binding
“The compendial methods cover a comprehensive
range of degradation forms which take into
account the knowledge of the molecular characteristics
up to this point in time. However, … these methods
may not be sufficient to appropriately detect the
thioether variant, even if it is present at high levels,
and that further powerful methods need to be
employed.”
London July 2, 2009
Source: M. Lispi et al. (2009) J. Pharm. Sci., DOI 10.1002/jps
G.-B. Kresse / CMC
27